Summary: The goal of the current project is to conduct an animal protection study with several VIG preparations in order to validate the in vitro neutralization assay. The results will greatly enhance the ability of CBER to streamline the testing of old and new VIG products and to alleviate the current shortage of VIG. These efforts will enhance CBER contributions to the Bioterrorism prevention efforts and will promote the testing of new recombinant vaccinia-based vaccines against HIV-1 and cancer. The specific goals are: Stage 1 To establish the optimal dose of vaccinia strains vSC56 and Wyeth that can be used to validate our in vitro viral neutralization assay. We expect death to occur within 10-20 days. Stage 2 and 3 To test multiple preparations of Vaccinia Immunoglobulins (VIG) and determine their IC50. This will involve determining the concentration of VIG required to protect 50 percent of the animals from a lethal dose of vaccinia virus infection. Results: - SCID mice were found to be more susceptible to vaccinia infection than nude mice.The immunodeficiency of SCID mice is more severe than that of the nude mice. - Both strains of vaccinia that were tested, Wyeth vaccine strain and vSC56, killled SCID mice between 25-35 days post infection (10^6 pfu/mouse, IP). - In the first in vivo mouse protection study, Baxter and MPH VIG were incubated with Wyeth vaccinia virus, either undiluted or at 1:10 and 1:100 dilutions of the VIG preparations. - Undiluted MPH VIG was completely protective. 1:10 and 1:100 dilutions were partially protective. They postponed animal death by 1-2 weeks. - The Baxter VIG (4oC lot) was less protective than the MPH. one death was observed with undiluted preparation and 1:10 VIG dilution. The 1:100 dilution was not significantly protective. - Since the concentration of IgG in Baxter VIG is >3 fold higher than in the MPH prpeparation, our preliminary results suggest that the MPH preparation should be the product of choice in the case of smallpox exposure or in the case of viremia associated with recombinant vaccinia vector vaccinations. On going studies: - Repeat of the first study using the Wyeth vaccinia virus and multiple dilutions of Baxter and MPH VIG preparations (1:4 serial dilutions). - Similar protection study using vSC56 virus (the virus strain which is used in the in vitro neutralization assays). - Testing of VIG subclasses in in vivo protection. - Testing of IVIG lots that were shown to have high vaccinia neutralization titers in the in vitro assay for their capacity to protect SCID mice from vaccinia infection.